Mutations in the Catalytic Subunit of Class IA PI3K Are Leukemogenic In Vivo.

The phosphoinositide 3-kinase (PI3K)-AKT pathway is a relevant signal transduction axis which regulates survival, growth and proliferation of hematopoietic cells through a variety of downstream targets. Constitutive activation of the PI3K-AKT pathway is observed in up to 70% of acute myelogenous leukemia (AML), primarily due to activating mutations in receptor tyrosine kinases (e.g. c-KIT) or RAS and is also present in T-cell-non-Hodgkins-lymphoma, mantle cell lymphoma, diffuse large B-cell lymphoma, Hodgkins Lymphoma and myelodysplastic syndrome (MDS). We analyzed the effect of activating point mutations in the catalytic subunit p110alpha of class IA PI3K (PIK3CA) in hematopoietic cells. We transfected early hematopoietic, IL-3 dependent cells (Ba/F3 cell line) with point mutations in the helical (exon 9, E542K, E545A) and kinase domain (exon 20, H1047R) of the p110alpha catalytic subunit and determined the effect of PI3K-AKT pathway activation on cellular proliferation, survival and leukemogenic expansion in vitro and in vivo. We demonstrate that the p110alpha point mutations constitutively activated the PI3K-AKT pathway and resulted in factor-independent growth of Ba/F3 cells. Proliferation and survival of the cells were inhibited in a time- and dose-dependent manner using inhibitors of either PI3K or AKT. The mammalian target of rapamycin (mTOR) was demonstrated to be important for mitogenic, but not anti-apoptotic signaling of mutant PIK3CA. In a syngenic mouse model, hematopoietic cells expressing mutant p110alpha induced a leukemia-like disease. FACS analysis demonstrated a median chimerism of 68% in the bone marrow and 35% in peripheral blood of diseased mice, which were characterized by anemia, leukemic infiltration of hematopoietic organs, and 90% mortality within 5 weeks. No substantial differences were observed between E542K, E545A and H1047R. Mice, which were transplanted with an activating c-KIT point mutation (D814V), showed a significantly reduced survival and 100% died within 9 days. In conclusion our data show, that PIK3CA point mutations, by activating the PI3K-AKT pathway, confer factor-independence to hematopoietic cells in vitro and induce leukemogenic activity in vivo. As mutant c-KIT (D814V) showed enhanced leukemogenic potential, signaling through c-KIT may involve additional downstream pathways which cooperate with activated PI3K in leukemia progression. Our model is useful for the differential investigation of the leukemogenic relevance of the PI3K-AKT pathway and pharmacological inhibitor studies.